Abstract: Soil microorganisms are important bioindicators of ecosystem health and functional changes. In this study, high-throughput sequencing and real-time quantitative PCR were employed to investigate the distribution characteristics, community structure, and gene abundance of carbon-fixing microorganisms in the mangrove wetlands of Hainan Dongzhai National Nature Reserve. Moreover, the disparities in carbon-fixing microbial communities between these zones and the key influencing factors were analyzed, aiming to provide a scientific basis for understanding the soil carbon fixation function of mangrove wetlands in tropical regions. The results showed that the dominant phyla of soil carbon-fixing bacteria in the soil were Pseudomonadota and Actinomycetota. The dominant genus in the bare mudflat area was Rubrivivax (12.93%), while the dominant genera in the mangrove area were Rubrivivax (10.57%) and Thiodictyon (10.68%), mainly composed of chemoheterotrophs and chemolithoautotrophs. The gene abundance of carbon-fixing bacteria was higher in bare mudflats than in the mangrove areas. According to alpha diversity indices (Chao1, Observed species, and Shannon), the bare mudflats farther from the ocean showed lower values than mangrove areas, while the reverse trend was observed in those closer to the ocean. Principal Coordinates Analysis (PCoA), Permutational Multivariate Analysis of Variance (PERMANOVA), and Analysis of Similarities (ANOSIM analyses) revealed significant differences in the soil carbon-fixing microbial community structures between bare mudflats and mangrove areas. Redundance analysis (RDA) further indicated that environmental factors such as TP (total phosphorus), SOC (soil organic carbon), SWC (soil water content), pH, TN (total nitrogen), and ROC (readily oxidizable carbon) had significant effects on the community structure of soil carbon-fixing microorganisms in the mangrove wetlands.